PROJECT SUMMARY Over the past two decades, one of the most exciting breakthroughs in biomedical research was the discovery of anti-retroviral therapy (ART), which curbs active HIV replication to nearly undetectable levels. Despite this great achievement, ART fails to cure HIV infection because latent reservoirs persist in tissues containing resting CD4 lymphocytes and other cell types that express low levels of viral products and can be reactivated in response to immune stimulation. Thus, these latent reservoirs are a critical barrier for a functional cure, making the need for new approaches even more urgent. To achieve this major goal, we must first identify novel, critical host factors that can be targeted therapeutically before we can leverage this knowledge for clinical intervention. To this end, this proposal will investigate a master host factor (KAP1) implicated in transducing a variety of physiologic immune signal inputs into proviral transcriptional outputs to reveal the molecular mechanisms and evaluate therapeutic potential. Our work is different from previous studies because it will help uncouple the contribution of the three major features influencing proviral transcription and fate (complex transcriptional circuit architecture, integration landscape, and immune cell state) to help devise rationale approaches for HIV eradication. We hypothesize that both the integration landscape and immune cell state regulate the outcome of an infection (active, latent, reactivated) through the function of critical master host factors like KAP1. New technologies and experimental approaches for investigating this problem provide unique opportunities to test this hypothesis and define the underlying molecular mechanisms. Specifically, we will investigate: 1) how KAP1 communicates with other host co-factors to assemble a positive transcription complex to promote the latency-reactivation switch in response to immune stimulation, 2) how the integration landscape and cell state influence KAP1-mediated proviral transcription activation and clonal expansion, 3) how KAP1 recognizes proviral chromatin to function as a ?scaffold? to couple transcription initiation and elongation, and 4) how KAP1 enzymatic functions help relieve the repression imparted by novel transcriptional co-repressors assembled at the provirus during the latency-reactivation switch. The knowledge generated in this study will establish the groundwork for interfering with the function of various ?druggable pockets? (e.g., enzymatic and chromatin reader domains) for HIV eradication strategies. The combination of high significance, innovation and use of a diverse set of experimental approaches in physiologically relevant systems (primary models of latency and patient samples) make this proposal unique. Collectively, these studies could have a revolutionary impact on our understanding of HIV latency biology and have therapeutic implications.